APOBEC3G (A3G) and related APOBEC3F (A3F) have broad anti-viral activity against retroviruses and hepatitis B virus (HBV). However, their regulation in viral natural target cells such CD4+ T lymphocytes, macrophages, and primary liver cells have not been well-studied. Our long-term goal is to elucidate the role of A3G and ASF in IFN-mediated innate immunity. The specific hypothesis is that certain human cells may utilize a novel IFN-mediated signaling pathway to upregulate the A3G and ASF proteins in the defense against viruses. We base the hypothesis on the following observations. 1) We showed that A3G was upregulated by interferons (IFN) in a cell-type dependent manner. IFN-a induced A3G in macrophages, primary hepatocytes and liver cell lines. IFN-y also induced A3G in liver cells. Neither cytokine could upregulate A3G in primary CD4+ T cells, while other known IFN-stimulated genes (ISG) such as PKR and IRF-1 were induced in all three cell-types. 2) In macrophages and liver cell lines, IFN-a induction of A3G was dependent upon JAK kinases. While the canonical IFN-a JAK/STAT signaling pathway requires both STAT1 and STAT2, we observed that induction of A3G and certain other ISG by IFN-a in HepSB liver cells was STAT2-dependent but STAT1-independent. The proposed experiments will focus on determining the nature of IFN-mediated A3G antiviral activity in macrophages and characterizing the specific molecular signaling pathway of IFN induction of A3G in macrophages and liver cells. The specific aims are: Aim 1) to study the role of IFN-a induced A3G and A3F expression in anti-HIV-1 activity of these proteins in macrophages. We will investigate the antiviral effect of A3G on HIV-1 in macrophages upon induction by IFN-a. The experiments will determine if IFN induction of A3G will result in functional antiviral activity in macrophages. Aim 2) to further characterize the STAT1-independent pathway of IFN-a induction of A3G and other ISG in liver cells and macrophages. IFN-a STAT1-independent pathways of gene transcription have not been described. These experiments will define components involved in the transcriptional activation of A3G and ASF. While we have determined that JAK kinases are required to induce A3G, we will further characterize the involvement of other STAT family proteins as well as potential novel mediators. The proposed research should provide insight into the antiviral role that A3G and ASF play both in natural immunity and in response to IFN treatment. Furthermore, the work would potentially enhance knowledge of general IFN-mediated signaling pathways. [unreadable] [unreadable] [unreadable]